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Cannabidiol (CBD), a major non-psychoactive phytocannabinoid in cannabis, is an effective treatment for some forms of epilepsy and pain. At high concentrations, CBD interacts with a huge variety of proteins, but which targets are most relevant for clinical actions is still unclear. Here we show that CBD interacts with Na v 1.7 channels at sub-micromolar concentrations in a state-dependent manner. Electrophysiological experiments show that CBD binds to the inactivated state of Na v 1.7 channels with a dissociation constant of about 50 nM. The cryo-EM structure of CBD bound to Na v 1.7 channels reveals two distinct binding sites. One is in the IV-I fenestration near the upper pore. The other binding site is directly next to the inactivated “wedged” position of the Ile/Phe/Met (IFM) motif on the short linker between repeats III and IV, which mediates fast inactivation. Consistent with producing a direct stabilization of the inactivated state, mutating residues in this binding site greatly reduced state-dependent binding of CBD. The identification of this binding site may enable design of compounds with improved properties compared to CBD itself. Cannabidiol (CBD), the nonpsychoactive component in cannabis, is an effective treatment for epilepsy and pain. Here, authors explored the mode of action of CBD on hNa v 1.7 channels through two distinct binding sites, suggesting a direct stabilization of the inactivated state of channels.

A highlight of the knowledge derived in large part from structural work on physical motions and chemical interactions involved in voltage sensing, pore opening, ion conductance and selectivity, and voltage-dependent inactivation mechanisms of the voltage-gated channels Na V and Ca V . Electrical signals generated by minute currents of ions moving across cell membranes are central to all rapid processes in biology. Initiation and propagation of electrical signals requires voltage-gated sodium (Na V ) and calcium (Ca V ) channels. These channels contain a tetramer of membrane-bound subunits or domains comprising a voltage sensor and a pore module. Voltage-dependent activation occurs as membrane depolarization drives outward movements of positive gating changes in the voltage sensor via a sliding-helix mechanism, which leads to a conformational change in the pore module that opens its intracellular activation gate. A unique negatively charged site in the selectivity filter conducts hydrated Na + or Ca 2+ rapidly and selectively. Ion conductance is terminated by voltage-dependent inactivation, which causes asymmetric pore collapse. This Review focuses on recent advances in structure and function of Na V and Ca V channels that expand our current understanding of the chemical basis for electrical signaling mechanisms conserved from bacteria to humans.

Photoactivatable drugs targeting ligand-gated ion channels open up new opportunities for light-guided therapeutic interventions. Photoactivable toxins targeting ion channels have the potential to control excitable cell activities with low invasiveness and high spatiotemporal precision. As proof-of-concept, we develop HwTxIV-Nvoc, a UV light-cleavable and photoactivatable peptide that targets voltage-gated sodium (Na V ) channels and validate its activity in vitro in HEK293 cells, ex vivo in brain slices and in vivo on mice neuromuscular junctions. We find that HwTxIV-Nvoc enables precise spatiotemporal control of neuronal Na V channel function under all conditions tested. By creating multiple photoactivatable toxins, we demonstrate the broad applicability of this toxin-photoactivation technology. Photoactivable toxins targeting ion channels have great potential to control cell activity. Here the authors report HwTxIV-Nvoc, a UV light-cleavable and photoactivatable peptide that targets voltage-gated sodium channels; they validate this in cells, brain slices and in vivo on mice neuromuscular junctions.

Targeted protein degradation/downregulation (TPD/TPDR) is a disruptive paradigm for developing therapeutics. <2% of ~600 E3 ligases have been exploited for this modality, and efficacy for multi-subunit ion channels has not been demonstrated. NEDD4-2 E3 ligase regulates myriad ion channels, but its utility for TPD/TPDR is uncertain due to complex regulatory mechanisms. Here, we identify a nanobody that binds NEDD4-2 HECT domain without disrupting catalysis sites as revealed by cryo-electron microscopy and in vitro ubiquitination assays. Recruiting NEDD4-2 to diverse ion channels (Ca V 2.2; KCNQ1; and epithelial Na + channel, ENaC, with a Liddle syndrome mutation) using divalent nanobodies (DiVas) strongly suppresses their surface density and function. Global proteomics indicates DiVa recruitment of endogenous NEDD4-2 to KCNQ1-YFP yields dramatically lower off-target effects compared to NEDD4-2 overexpression. The results establish utility of NEDD4-2 recruitment for TPD/TPDR, validate ion channels as susceptible to this modality, and introduce a general method to generate ion channel inhibitors. Researchers develop a new way to selectively remove ion channel proteins by recruiting the body’s own NEDD4-2 enzyme using custom nanobodies, offering a precise and general strategy for future drug development.

Voltage-gated sodium channels (VGSC) are transmembrane proteins that generate an action potential in excitable cells and play an essential role in neuronal signaling. Since VGSCs play a crucial role in nerve transmission they have become primary targets for a broad range of commercial insecticides. RNA interference (RNAi) is a valuable reverse genetics tool used in functional genomics, but recently, it has also shown promise as a novel agent that could be used to control agricultural insect pests. In this study, we targeted the VGSC ( MpNa v ) gene in the peach-potato aphid Myzus persicae , by oral feeding of artificial diets mixed with dsRNAs. Knock-down of MpNa v gene expression caused up to 65% mortality in 3 rd instar nymphs. Moreover, significantly lower fecundity and longevity was observed in adult aphids that had been fed with dsMpNa v solution at the nymphal stage. Analysis of gene expression by qRT-PCR indicated that the aphid mortality rates and the lowered fecundity and longevity were attributable to the down-regulation of MpNa v by RNAi. Taken together, our results show that MpNa v is a viable candidate target gene for the development of an RNAi-based bio-aphicide.

Here we report the pharmacologic blockade of voltage-gated sodium ion channels (Na V s) by a synthetic saxitoxin derivative affixed to a photocleavable protecting group. We demonstrate that a functionalized saxitoxin (STX-eac) enables exquisite spatiotemporal control of Na V s to interrupt action potentials in dissociated neurons and nerve fiber bundles. The photo-uncaged inhibitor (STX-ea) is a nanomolar potent, reversible binder of Na V s. We use STX-eac to reveal differential susceptibility of myelinated and unmyelinated axons in the corpus callosum to Na V -dependent alterations in action potential propagation, with unmyelinated axons preferentially showing reduced action potential fidelity under conditions of partial Na V block. These results validate STX-eac as a high precision tool for robust photocontrol of neuronal excitability and action potential generation. Photocaged molecules have advantages in terms of temporal and spatial control compared to conventional pharmacological compounds. The authors present a synthetic saxitoxin derivative affixed to a photocleavable group for precise modulation of Na channels.

Key Points Renal perfusion pressure, glomerular filtration and net renal sodium reabsorption maintain volume homeostasis Cell stiffness and myogenic tone contribute to peripheral vascular resistance in the presence of aldosterone and normal sodium concentrations in the arterial circulation Structural similarities exist between the amiloride-sensitive sodium channels in the epithelial and endothelial cells Distinctions based on functional differences, especially in response to changes in extracellular sodium concentrations, might permit development of novel pharmacological approaches to differentially target epithelial and endothelial sodium channels Agents that preferentially inhibit the vascular channels could lower systemic blood pressure without the associated inhibition of potassium secretion that accompanies the available sodium channel blockers and mineralocorticoid receptor antagonists Blood pressure control is influenced by amiloride-sensitive sodium channels in the vascular and epithelial homeostatic systems in the kidney. Here, the authors describe the expression and regulation of sodium channels, and they outline the emerging evidence that differences between sodium channel complexes expressed in the epithelia and endothelia might permit novel therapeutic approaches to lower systemic blood pressure without the adverse effects associated with the sodium channel blockers that are currently available. Sodium transport in the distal nephron is mediated by epithelial sodium channel activity. Proteolytic processing of external domains and inhibition with increased sodium concentrations are important regulatory features of epithelial sodium channel complexes expressed in the distal nephron. By contrast, sodium channels expressed in the vascular system are activated by increased external sodium concentrations, which results in changes in the mechanical properties and function of endothelial cells. Mechanosensitivity and shear stress affect both epithelial and vascular sodium channel activity. Guyton's hypothesis stated that blood pressure control is critically dependent on vascular tone and fluid handling by the kidney. The synergistic effects, and complementary regulation, of the epithelial and vascular systems are consistent with the Guytonian model of volume and blood pressure regulation, and probably reflect sequential evolution of the two systems. The integration of vascular tone, renal perfusion and regulation of renal sodium reabsorption is the central underpinning of the Guytonian model. In this Review, we focus on the expression and regulation of sodium channels, and we outline the emerging evidence that describes the central role of amiloride-sensitive sodium channels in the efferent (vascular) and afferent (epithelial) arms of this homeostatic system.

Salty taste is one of the five basic tastes and is often elicited by NaCl. Because excess sodium intake is associated with many health problems, it could be useful to have salt taste enhancers that are not sodium based. In this study, the regulation of NaCl-induced responses was investigated in cultured human fungiform taste papillae (HBO) cells with five arginyl dipeptides: Ala-Arg (AR), Arg-Ala (RA), Arg-Pro (RP), Arg-Glu (RE), and Glu-Arg (ER); and two non-arginyl dipeptides: Asp-Asp (DD) and Glu-Asp (ED). AR, RA, and RP significantly increased the number of cell responses to NaCl, whereas no effect was observed with RE, ER, DD, or ED. We also found no effects with alanine, arginine, or a mixture of both amino acids. Pharmacological studies showed that AR significantly increased responses of amiloride-sensitive but not amiloride-insensitive cells. In studies using small interfering RNAs (siRNAs), responses to AR were significantly decreased in cells transfected with siRNAs against epithelial sodium channel ENaCα or ENaCδ compared to untransfected cells. AR dramatically increased NaCl-elicited responses in cells transfected with NHE1 siRNA but not in those transfected with ENaCα or ENaCδ siRNAs. Altogether, AR increased responses of amiloride-sensitive cells required ENaCα and ENaCδ.

Ion channels are essential for various physiological processes, and their defects are associated with many diseases. Previous research has revealed that a Terahertz electromagnetic field can alter the channel conductance by affecting the motion of chemical groups of ion channels, and hence regulate the electric signals of neurons. In this study, we conducted molecular dynamics simulations to systematically investigate the effects of terahertz electromagnetic fields on the ion permeation of voltage-gated potassium and sodium channels, particularly focusing on the bound ions in the selectivity filters that have not been extensively studied previously. Our results identified multiple new characteristic frequencies and showed that 1.4, 2.2, or 2.9 THz field increases the ion permeability of K v 1.2, and 2.5 or 48.6 THz field increases the ion permeability of Na v 1.5. Such effects are specific to the frequencies and directions of the electric field, which are determined by the intrinsic oscillation motions of the permeating ions in the selectivity filter or certain chemical groups of the ion channels. The amplitude of the THz field positively correlates with the change in ion permeation. This study demonstrates that THz fields can specifically regulate ion channel conductances by multiple mechanisms, which may carry great potential in biomedical applications. Terahertz (THz) electromagnetic fields can alter ion channel conductance, however, the mechanism by which they do so is not fully understood. Here, the authors perform molecular dynamics simulations to reveal that THz waves with specific frequencies can enhance the ion permeation by directly affecting the motion of K + or Na + bound at the selectivity filter of the K v 1.2 and Na v 1.5 channels.